Acute myelogenous leukemia (AML) is characterized by the excessive proliferation and abnormal differentiation of immature myeloid progenitor cells. The long term goal of this proposal is to identify the genes which transform myeloid cells with an emphasis on those that alter normal differentiation. Our approach utilizes a series of tumorigenic, IL-3- dependent myeloid cell lines, isolated from retrovirus induced leukemias. These transformed cells do not terminally differentiate but are unaltered in their growth factor requirements. To identify the genes associated with this phenotype of transformation, we looked for rearrangements of known proto-oncogenes or for common sites of proviral integration representing putative proto-oncogene loci. These studies have identified two genetically linked genomic regions of proviral integrations (Evi-1 and CB- 1/Fim-3), which are commonly rearranged. Viral integrations in these sites are associated with the activation of the expression of a gene encoding a protein predicted to be a member of the zinc finger family of transcriptional regulatory proteins. We hypothesize that the aberrant expression of this gene in myeloid cells alters their differentiation program by affecting the transcription of genes which are required for differentiation. The overall objective of this proposal is to determine the role of the Evi-1 gene in myeloid transformation. We will determine whether the Evi-1 gene codes for a nuclear, DNA binding protein that is involved in transcriptional regulation. Its role in transformation will be assessed by using retroviral vectors to introduce the gene in normal myeloid stem cells. The normal function and transforming potential of the gene will be examined by developing transgenic mice expressing the Evi-1 gene and by using homologous recombination in ES cells to develop murine strains which lack the gene. The mechanisms of Evi-1 activation by viral integration in the CB-1/Fim-3 locus will be determined. The human gene will be characterized and its potential role in AML and other malignancies will be examined. Lastly, we will continue to characterize integration sites in IL-3-dependent myeloid cell lines to identify novel genes involved in their transformation. These studies will provide new insights into the genes involved in myeloid transformation and the genes and mechanisms involved in normal hematopoietic differentiation.